1. Field of the Invention
The invention relates to a process for the preparation of purple membrane containing bacteriorhodopsin.
2. The Prior Art
Bacteriorhodopsin is a chromoprotein which is formed by halobacteria, such as Halobacterium halobium. It consists of a protein moiety (bacterioopsin) and a chromophore (retinal), and it occurs in the cell membrane of halobacteria in the form of the so-called purple membrane (hereinafter also called PM). In the PM, bacteriorhodopsin is arranged in a two-dimensional crystalline, hexagonal lattice. By virtue of its physical properties, bacteriorhodopsin might be employed in biocomputers (cf. Biosystems 1986, 19, 223-236) or as a holographic storage material in optical data processing (cf. Biophysics 1985, 30(5), 962-967). For this purpose, however, it is necessary to be able to obtain large amounts of purple membranes. Since the physical differences between purple membranes and contaminating membrane fragments which do not contain bacteriorhodopsin are small, it is difficult to separate the purple membrane from the contaminating fragments.
According to D. Oesterhelt, W. Stoeckenius, Nature 1971, 233, 149-154, and Methods in Enzymology 1974, 31, 667-678, it is known to carry out the preparation by centrifugation of the purple membrane in a sucrose density gradient from 0.5 to 1.5M or from 30 to 50% by weight sucrose. B. M. Becher, J. Y. Cassim, Preparative Biochemistry 1975, 5(2), 161-178, describe a centrifugation of the purple membrane by means of a stepped sucrose gradient having steps of 45, 40, 38, 36 and 16% by weight of sucrose.
Due to the complicated apparatus in these gradient centrifugations, which, for example, require ultracentrifuges, a variable scale-up with regard to the amounts of bacteriorhodopsin to be obtained is not possible, which means that only comparatively low amounts of PM are isolated in one work-up.
Furthermore, the preparation of the sucrose gradients and the removal of the employed sucrose from the PM obtained are procedure steps which involve large amounts of time.